High-fat diet exacerbates renal dysfunction in SHR: reversal by induction of HO-1-adiponectin axis

High-dietary fat intake is a major risk factor for development of metabolic and cardiovascular-renal dysfunction including obesity, coronary artery disease, hypertension, and chronic renal failure. We examined the effect of a high-fat diet on renal function and morphology in spontaneously hypertensive rats (SHR), a phenotype designed to mimic metabolic syndrome. High-fat diet induced increase (P < 0.05) in blood pressure, body weight, and renal lipid deposition in these rats. This increase in body weight was accompanied by elevations (P < 0.05) of blood glucose and low-density lipoprotein (LDL) levels, a decrease (P < 0.05) in adiponectin and increases (P < 0.05) in plasma monocyte chemotactic protein-1 (MCP-1) along with renal macrophage infiltration. These pathophysiological perturbations were attenuated (P < 0.05) by heme oxygenase-1 (HO-1) induction by treatment with cobalt protoporphyrin (CoPP). Further effects of CoPP included increased (P < 0.05) renal expression of adiponectin along with enhancement (P < 0.05) of pAKT, pAMPK, and p-eNOS in SHRs fed a high-fat diet. Prevention of such beneficial effects of CoPP by the concurrent administration of the heme-HO inhibitor stannous mesoporphyrin (SnMP) corroborates the role of HO system in mediating such effects. Taken together, our results demonstrate that high-fat diet induces a metabolic syndrome-like phenotype in hypertensive rats, which is amenable to rescue by increases in HO-1- and adiponectin-dependent pathways.     

Sensitization of Candida albicans biofilms to various antifungal drugs by cyclosporine A

Background

Biofilms formed by Candida albicans are resistant towards most of the available antifungal drugs. Therefore, infections associated with Candida biofilms are considered as a threat to immunocompromised patients. Combinatorial drug therapy may be a good strategy to combat C. albicans biofilms.

Methods

Combinations of five antifungal drugs- fluconazole (FLC), voriconazole (VOR), caspofungin (CSP), amphotericin B (AmB) and nystatin (NYT) with cyclosporine A (CSA) were tested in vitro against planktonic and biofilm growth of C. albicans. Standard broth micro dilution method was used to study planktonic growth, while biofilms were studied in an in vitro biofilm model. A chequerboard format was used to determine fractional inhibitory concentration indices (FICI) of combination effects. Biofilm growth was analyzed using XTT-metabolic assay.

Results

MICs of various antifungal drugs for planktonic growth of C. albicans were lowered in combination with CSA by 2 to 16 fold. Activity against biofilm development with FIC indices of 0.26, 0.28, 0.31 and 0.25 indicated synergistic interactions between FLC-CSA, VOR-CSA, CSP-CSA and AmB-CSA, respectively. Increase in efficacy of the drugs FLC, VOR and CSP against mature biofilms after addition of 62.5 μg/ml of CSA was evident with FIC indices 0.06, 0.14 and 0.37, respectively.

Conclusions

The combinations with CSA resulted in increased susceptibility of biofilms to antifungal drugs. Combination of antifungal drugs with CSA would be an effective prophylactic and therapeutic strategy against biofilm associated C. albicans infections.     

Low plasma albumin levels are associated with increased plasma protein glycation and HbA1c in diabetes

Albumin is one of the most abundant plasma proteins and is heavily glycated in diabetes. In this study, we have addressed whether variation in the albumin levels influence glycation of plasma proteins and HbA1c. The study was performed in three systems: (1) streptozotocin (STZ)-induced diabetic mice plasma, (2) diabetic clinical plasma, and (3) in vitro glycated plasma. Diabetic mice and clinical plasma samples were categorized as diabetic high albumin plasma (DHAP) and diabetic low albumin plasma (DLAP) on the basis of their albumin levels. For the in vitro experiment, two albumin levels, high albumin plasma (HAP) and low albumin plasma (LAP), were created by differential depletion of plasma albumin. Protein glycation was studied by using a combination of two-dimensional electrophoresis (2DE), Western blotting, and LC-MS(E). In both mice and clinical experiments, an increased plasma protein glycation was observed in DLAP than in DHAP. Additionally, plasma albumin levels were negatively correlated with HbA1c. The in vitro experiment with differential depletion of albumin mechanistically showed that the low albumin levels are associated with increased plasma protein glycation and that albumin competes for glycation with other plasma proteins.     

Rapid,in situ detection of Agrobacterium tumefaciens attachment to leaf tissue

Attachment of the plant pathogen Agrobacterium tumefaciens to host plant cells is an early and necessary step in plant transformation and agroinfiltration processes. However, bacterial attachment behavior is not well understood in complex plant tissues. Here we developed an imaging‐based method to observe and quantify A. tumefaciens attached to leaf tissue in situ. Fluorescent labeling of bacteria with nucleic acid, protein, and vital dyes was investigated as a rapid alternative to generating recombinant strains expressing fluorescent proteins. Syto 16 green fluorescent nucleic acid stain was found to yield the greatest signal intensity in stained bacteria without affecting viability or infectivity. Stained bacteria retained the stain and were detectable over 72 h. To demonstrate in situ detection of attached bacteria, confocal fluorescent microscopy was used to image A. tumefaciens in sections of lettuce leaf tissue following vacuum‐infiltration with labeled bacteria. Bacterial signals were associated with plant cell surfaces, suggesting detection of bacteria attached to plant cells. Bacterial attachment to specific leaf tissues was in agreement with known leaf tissue competencies for transformation with Agrobacterium. Levels of bacteria attached to leaf cells were quantified over time post‐infiltration. Signals from stained bacteria were stable over the first 24 h following infiltration but decreased in intensity as bacteria multiplied in planta. Nucleic acid staining of A. tumefaciens followed by confocal microscopy of infected leaf tissue offers a rapid, in situ method for evaluating attachment of A. tumefaciens' to plant expression hosts and a tool to facilitate management of transient expression processes via agroinfiltration. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012     

Novel insights through the integration of structural and functional genomics data with protein networks

In recent years, major advances in genomics, proteomics, macromolecular structure determination, and the computational resources capable of processing and disseminating the large volumes of data generated by each have played major roles in advancing a more systems-oriented appreciation of biological organization. One product of systems biology has been the delineation of graph models for describing genome-wide protein-protein interaction networks. The network organization and topology which emerges in such models may be used to address fundamental questions in an array of cellular processes, as well as biological features intrinsic to the constituent proteins (or "nodes") themselves. However, graph models alone constitute an abstraction which neglects the underlying biological and physical reality that the network's nodes and edges are highly heterogeneous entities. Here, we explore some of the advantages of introducing a protein structural dimension to such models, as the marriage of conventional network representations with macromolecular structural data helps to place static node and edge constructs in a biologically more meaningful context. We emphasize that 3D protein structures constitute a valuable conceptual and predictive framework by discussing examples of the insights provided, such as enabling in silico predictions of protein-protein interactions, providing rational and compelling classification schemes for network elements, as well as revealing interesting intrinsic differences between distinct node types, such as disorder and evolutionary features, which may then be rationalized in light of their respective functions within networks.     

Regulation of exocytosis by the exocyst subunit Sec6 and the SM protein Sec1

Trafficking of protein and lipid cargo through the secretory pathway in eukaryotic cells is mediated by membrane-bound vesicles. Secretory vesicle targeting and fusion require a conserved multisubunit protein complex termed the exocyst, which has been implicated in specific tethering of vesicles to sites of polarized exocytosis. The exocyst is directly involved in regulating soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein receptor (SNARE) complexes and membrane fusion through interactions between the Sec6 subunit and the plasma membrane SNARE protein Sec9. Here we show another facet of Sec6 function-it directly binds Sec1, another SNARE regulator, but of the Sec1/Munc18 family. The Sec6-Sec1 interaction is exclusive of Sec6-Sec9 but compatible with Sec6-exocyst assembly. In contrast, the Sec6-exocyst interaction is incompatible with Sec6-Sec9. Therefore, upon vesicle arrival, Sec6 is proposed to release Sec9 in favor of Sec6-exocyst assembly and to simultaneously recruit Sec1 to sites of secretion for coordinated SNARE complex formation and membrane fusion.     

The association between Barrett's esophagus and Helicobacter pylori infection: a meta-analysis

Objective: The effect of Helicobacter pylori on Barrett’s esophagus is poorly understood. We conducted a meta‐analysis to summarize the existing literature examining the effect that H. pylori has on Barrett’s esophagus. Design: We performed a comprehensive search to identify studies pertaining to the association between H. pylori and Barrett’s esophagus. We conducted meta‐regression analyses to identify sources of variation in the effect of H. pylori on Barrett’s esophagus. Results: Our analysis included a total of 49 studies that examined the effect of H. pylori on Barrett’s esophagus and seven studies that examined the effect of cag A positivity on Barrett’s esophagus. Overall, H. pylori, and even more so cag A, tended to be protective for Barrett’s esophagus in most studies; however, there was obvious heterogeneity across studies. The effect of H. pylori on Barrett’s esophagus varied by geographic location and in the presence of selection and information biases. Only four studies were found without obvious selection and information bias, and these showed a protective effect of H. pylori on Barrett’s esophagus (Relative risk = 0.46 [95% CI: 0.35, 0.60]). Conclusions: Estimates for the effect of H. pylori on Barrett’s esophagus were heterogeneous across studies. We identified selection and information bias as potential sources of this heterogeneity. Few studies without obvious selection and information bias have been conducted to examine the effect of H. pylori on Barrett’s esophagus, but in these, H. pylori infection is associated with a reduced risk of Barrett’s esophagus.     

Single-cell census of mechanosensitive channels in living bacteria

Bacteria are subjected to a host of different environmental stresses. One such insult occurs when cells encounter changes in the osmolarity of the surrounding media resulting in an osmotic shock. In recent years, a great deal has been learned about mechanosensitive (MS) channels which are thought to provide osmoprotection in these circumstances by opening emergency release valves in response to membrane tension. However, even the most elementary physiological parameters such as the number of MS channels per cell, how MS channel expression levels influence the physiological response of the cells, and how this mean number of channels varies from cell to cell remain unanswered. In this paper, we make a detailed quantitative study of the expression of the mechanosensitive channel of large conductance (MscL) in different media and at various stages in the growth history of bacterial cultures. Using both quantitative fluorescence microscopy and quantitative Western blots our study complements earlier electrophysiology-based estimates and results in the following key insights: i) the mean number of channels per cell is much higher than previously estimated, ii) measurement of the single-cell distributions of such channels reveals marked variability from cell to cell and iii) the mean number of channels varies under different environmental conditions. The regulation of MscL expression displays rich behaviors that depend strongly on culturing conditions and stress factors, which may give clues to the physiological role of MscL. The number of stress-induced MscL channels and the associated variability have far reaching implications for the in vivo response of the channels and for modeling of this response. As shown by numerous biophysical models, both the number of such channels and their variability can impact many physiological processes including osmoprotection, channel gating probability, and channel clustering.